The Primary Study About Molecular Mechanism Of Lycorine Inhibiting HL-60 Cell Proliferation Via Up-regulating P21 Gene Expression

Leukemia, one of the most threatening hematological malignant cancer today, has been found very sensitive to anti-cancer chemotherapeutic reagents which either interfere with cell cycles or cause apoptosis. Lycorine is one member of Amaryllidaceae family, showing a natural anti-cancer activity. We used human acute promyelocytic leukemia cell line HL-60 to study the anti-tumor mechanisms of lycorine in two aspects: apoptosis and cell cycle arrest.Part 1 Gene chip detected the differences in transcription levels of apoptosis and proliferation associated genes in HL-60 cell which is divided into 2 groups and treated by different concentration of lycorineThe cells were incubated with 0 or 5.0μM lycorine for 24 h. The group treated without lycorine served as controls. The total RNA was extracted from two groups. Gene chip was used to detect the differences in transcription levels of apoptosis or proliferation associated genes between two groups. The gene chip included 494 genes which associated with apoptosis and proliferation. 91 genes were detected with significant changes of expression levels in total. 79 genes were up-regulated, meanwhile 12 genes were down-regulated. Some important genes were found with different transcription levels between 2 groups, such as p21,Caspase-7,DAPK1,TNF-αand Cyclin G1. Their changes in transcription levels would cause cell apoptosis and cell cycle arrest. So lycorine could induce changes in transcription levels of some key genes, these changes would cause cell apoptosis and cell cycle arrest.Part 2 The primary mechanisms study of lycorine inhibiting HL-60 cell proliferationFirstly, we used RT-PCR to detect the gene chip result about p21 gene, and the result of RT-PCR was consistent with the gene chip. Then we used Western blot to detect the expression changes of p21 protein and its' downstream protein Cdc2, Cdk2 and Cyclin E after HL-60 cells were incubated with lycorine. The cells were divided into 4 groups and incubated with 0, 1.25, 2.5 and 5.0μM lycorine for 24 h. The groups treated without lycorine served as controls. As a result, in 0, 1.25, 2.5 and 5.0μM groups, the expression of p21 protein was up-regulated, meanwhile the expression of Cdc2, Cdk2 and Cyclin E was down-regulated. So the lycorine could interfere with the cell cycle signal pathway in HL-60 cell, and induce cell cycle arrest.